Insulin resistance and beta cell failure play key roles in the pathogenesis of diabetes. Targeted mutagenesis in mice suggests that both abnormalities can be ascribed to impaired insulin signaling. Over the past funding cycle, the PI has endeavored to meet three goals: i, to define the contribution of different insulin target tissues to the pathogenesis of insulin resistance;ii, to understand whether different IRS proteins play separate or overlapping roles in insulin action;and Hi, to examine the role of insulin and IGF signaling in pancreatic beta cells to the pathogenesis of beta cell failure. These studies have led to a reassessment of the site(s) of onset of insulin resistance, by emphasizing the contribution to this process by tissues that do not display insulin dependent glucose uptake, such brain and liver. The studies have also contributed to establishing the idea that pancreatic beta cells are an "insulin-sensitive" cell type. Building on these lessons, the PI proposes studies of the integrated physiology of insulin action that will focus primarily on the contribution of liver, brain and pancreatic betaa cells to insulin resistance. These studies will be carried out using a genetic approach in which mutations affecting insulin signaling are introduced in mice and the consequences analyzed by thorough metabolic phenotyping. There are three aims in this proposal. In Aim 1, the PI will analyze the role of different organs in insulin action. Studies during the past cycle have established that insulin signaling in liver, as well as atypical target tissues, such as brain and pancreatic acells, is sufficient to restore metabolic control in insulin receptor (InsR)-deficient mice. The PI now wishes to test whether restoring InsR function at sites of insulin-dependent glucose uptake and disposal, such as skeletal muscle and adipocytes, would also have a similar effect. This question will be addressed using a locus knock-in approach to express InsR in mice. In Aim 2, the PI will use transgenic knockout mice expressing InsR solely in brain, liver and beta cells to study the role of direct and indirect mechanisms in insulin control of hepatic glucose production. To this end, the PI will employ conditional tissue knock-ins to restore InsR function at additional sites, with an emphasis on the arcuate and paraventricular hypothalamic nuclei. In Aim 3, the PI will study the role of insulin resistance in the regulation of beta cell mass. The PI will perform lineage tracing experiments of Neurogenin3-expressing endocrine progenitor cells in insulin-resistant mice and introduce mutations affecting insulin/IGF signaling in the same cell type to study the function of these pathways in endocrine cell differentiation.